Yang Yi, Mao Yongjiang, Kelly Patrick, Yang Zhangpin, Luan Lu, Zhang Jilei, Li Jing, El-Mahallawy Heba S, Wang Chengming
Jiangsu Co-innovation Center for the Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou University College of Veterinary Medicine, Yangzhou 225009, Jiangsu, China.
Parasit Vectors. 2014 Aug 31;7:413. doi: 10.1186/1756-3305-7-413.
Theileria spp. are tick transmitted protozoa that can infect large and small ruminants causing disease and economic losses. Diagnosis of infections is often challenging, as parasites can be difficult to detect and identify microscopically and serology is unreliable. While there are PCR assays which can identify certain Theileria spp., there is no one PCR that has been designed to identify all recognized species that occur in ruminants and which will greatly simplify the laboratory diagnoses of infections.
Primers and probes for a genus-specific pan-Theileria FRET-qPCR were selected by comparing sequences of recognized Theileria spp. in GenBank and the test validated using reference organisms. The assay was also tested on whole blood samples from large and small ruminants from nine provinces in China.
The pan-Theileria FRET-qPCR detected all recognized species but none of the closely related protozoa. In whole blood samples from animals in China, Theileria spp. DNA was detected in 53.2% of the sheep tested (59/111), 44.4% of the goats (120/270) and 30.8% of the cattle (380/1,235). Water buffaloes (n = 29) were negative. Sequencing of some of the PCR products showed cattle in China were infected with T. orientalis/T. sergenti/T. buffeli group while T. ovis and T. luwenshuni were found in sheep and T. luwenshuni in goats. The prevalence of Theileria DNA was significantly higher in Bos p. indicus than in Bos p. taurus (77.7% vs. 18.3%) and copy numbers were also significantly higher (10(4.88) vs. 10(3.00) Theileria 18S rRNA gene copies/per ml whole blood).
The pan-Theileria FRET-qPCR can detect all recognized Theileria spp. of ruminants in a single reaction. Large and small ruminants in China are commonly infected with a variety of Theileria spp.
泰勒虫属是通过蜱传播的原生动物，可感染大型和小型反刍动物，导致疾病和经济损失。感染的诊断通常具有挑战性，因为寄生虫在显微镜下难以检测和识别，血清学也不可靠。虽然有一些聚合酶链反应（PCR）检测方法可以识别某些泰勒虫属，但没有一种PCR检测方法是专门设计用于识别反刍动物中所有已知物种的，这将大大简化感染的实验室诊断。
通过比较GenBank中已知泰勒虫属的序列，选择了一种属特异性泛泰勒虫荧光共振能量转移定量PCR（FRET-qPCR）的引物和探针，并使用参考生物体对该检测方法进行了验证。该检测方法还在中国九个省份的大型和小型反刍动物的全血样本上进行了测试。
泛泰勒虫FRET-qPCR检测到了所有已知物种，但未检测到任何密切相关的原生动物。在中国动物的全血样本中，53.2%的测试绵羊（59/111）、44.4%的山羊（120/270）和30.8%的牛（380/1235）检测到泰勒虫属DNA。水牛（n = 29）检测结果为阴性。对一些PCR产物进行测序表明，中国的牛感染了东方泰勒虫/瑟氏泰勒虫/ buffeli组，而绵羊中发现了卵形泰勒虫和吕氏泰勒虫，山羊中发现了吕氏泰勒虫。瘤牛中泰勒虫DNA的流行率显著高于普通牛（77.7%对18.3%），拷贝数也显著更高（10（4.88）对10（3.00）泰勒虫18S rRNA基因拷贝数/毫升全血）。
泛泰勒虫FRET-qPCR可以在一次反应中检测到反刍动物中所有已知泰勒虫属物种。中国的大型和小型反刍动物普遍感染多种泰勒虫属物种。
